The small subunit ribosomal RNA (SrRNA) gene of Entamoeba gingivalis was amplified by PCR and the product of 1.9-kb long was cloned into a plasmid vector pUC18. Four clones were isolated and sequenced. The insert DNAs were 1918-to 1921-bp long and A+T rich (65.5%). The four SrRNA sequences of E.gingivalis were aligned with those of nine related protozoans searching for E.gingivalis-specific sequence. A sequence of 28 oligonucleotides was chosen, chemically synthesized, and labeled with digoxigenin for a DNA probe. The probe thus constructed was shown to hybridize only with either the SrRNA-coding DNAs or the cells of the two E.gingivalis strains and not with those of other protozoans and oral fungi tested. A representative SrRNA-sequence was analyzed and a phylogenetic tree was constructed by neighbor-joining (NJ) method. Among the protists examined, E.gingivalis was placed next to Entamoeba histolytica as expected from the traditional taxonomy.